Bioavailability
Pharmacokinetics is the study of the fate of pharmaceuticals from the time they are ingested until they are eliminated from the body. The sequence of events for an oral composition includes absorption through the various mucosal surfaces, distribution via the blood stream to various tissues, biotransformation in the liver and other tissues, action at the target site, and elimination of drug or metabolites in urine or bile.
Bioavailability of a drug (pharmaceutical composition) following oral dosing is a critical pharmacokinetic determinant which can be approximated by the following formula: EQU F.sub.oral =F.sub.ABS .times.F.sub.G .times.F.sub.H
F.sub.oral is oral bioavailability fraction, which is the fraction of the oral dose that reaches the circulation in an active, unchanged form. F.sub.oral is less than 100% of the active ingredient in the oral dose for three reasons: drug is not absorbed through the GI tract and is eliminated in the feces; drug is biotransformed by the cells of the intestine (to an inactive metabolite); or drug is eliminated by the cells of the liver, either by biotransformation and/or by transport into the bile. Thus, oral bioavailability is the product of the fraction of the oral dose that is absorbed (F.sub.ABS), the fraction of the absorbed dose that successfully reaches the blood side of the gastrointestinal tract (F.sub.G), and the fraction of the drug in the GI blood supply that reaches the heart side of the liver (F.sub.H). Previous drug formulations have attempted to increase drug efficacy by increasing drug absorption. For example, methods have been used to increase drug absorption using liposomes as carriers and designing more lipophilic drugs. These methods can increase drug absorption; however, they fail to address other ways of increasing drug bioavailability.
Liver Biotransformation and Biliary Secretion
The liver affects drug bioavailability. All blood from the gastrointestinal tract passes through the liver before going elsewhere in the body in all mammals, including humans. Due to its location, liver transformation of orally dosed drugs has a substantial "first-pass effect" on drug bioavailability that was thought to exceed effects in the gut, as discussed by Yun K. Tam in "Individual Variation in First-Pass Metabolism," Clin. Pharmacokinetics 25:300-328 (1993):
Enzyme activity in the small intestine is lower than in the liver. In humans, the liver to intestine cytochrome P450 ratio has been reported as .apprxeq.20, suggesting that the contribution of intestinal phase I biotransformation to the overall metabolism of a drug is unlikely to be important. (op. cit. 303)
Elimination of active drug by the liver occurs by one or both of two general pathways, namely biotransformation of the drug and excretion of the drug into the bile. Biotransformation reactions have been classified into two broadly defined phases. Phase I biotransformation often utilizes reactions catalyzed by the cytochrome P450 enzymes, which are manifold and active in the liver and transform many chemically diverse drugs. A second biotransformation phase can add a hydrophilic group, such as glutathione, glucuronic acid or sulfate, to increase water solubility and speed elimination through the kidneys.
Hepatocytes have contact with many types of blood and other fluid-transport vessels, such as the portal vein (nutrient and drug-rich blood from the gut), the hepatic arteries (oxygenated blood direct from the heart), the hepatic veins (efflux), lymphatics (lipids and lymphocytes), and bile ducts. The biliary ducts converge into the gall bladder and common bile duct that excretes bile into the upper intestine, aiding digestion. Bile also contains a variety of excretory products including hydrophobic drugs and drug metabolites.
Traditional solubility rate limiting approaches to increasing drug efficacy have focused on increasing solubility and membrane permeability. Where metabolism-based approaches have been considered, they have focused on biotransformation in liver. Although methods exist that affect biotransformation in the liver, these methods are inadequate because they affect general liver metabolism and can produce broad non-specific systemic effects.
Cytochromes
Most biotransformation is performed by enzymes called "mixed function oxidases" containing cytochromes, molecules with iron-containing rings, that help reduce oxygen to water. The cytochrome-containing enzymes that transform drugs use radical oxygen. When oxygen is reduced to its reactive radical form, it reacts immediately with the drug at the oxygen reduction site.
Most research on cytochromes involved in drug biotransformation focuses on inter-individual differences in cytochrome activity because such differences appear to be the dominant mechanism for differences in elimination of pharmaceuticals between individuals. Large inter-individual differences observed in the effects of drugs are at least in part determined by the variation of the expression and catalytic activity of the cytochromes P450.
The sources of the inter-individual variation in the catalytic activity of the cytochromes P450 can be divided into four general categories. The first is the influence of genetics on the expression of the cytochromes P450. Significant inter-individual variability can occur for each of the cytochromes P450. Genetic polymorphisms have been well characterized for the two cytochromes P450 responsible for debrisoquine/sparteine metabolism (CYP2D6; cytochrome families are defined below) and (S)-mephenytoin 4'-hydroxylation (possibly CYP2C19). The second source of inter-individual differences is that several of the human cytochromes P450 are inducible. That is, the content as well as the catalytic activity of these cytochromes P450 is increased by exposure of an individual to particular classes of drugs, endogenous compounds, and environmental agents. Thirdly, the activity of the cytochromes P450 can be inhibited or the cytochromes P450 can be inactivated by drugs and environmental compounds. This includes competitive inhibition between substrates of the same cytochrome P450, inhibition by agents that bind sites on the cytochrome P450 other than the active site, and suicidal inactivation of the cytochrome P450 by reactive intermediates formed during the metabolism of an agent. Another source of inter-individual differences is host factors. These factors include disease states, diet, and hormonal influences. Inter-individual differences in the level of expression and catalytic activity of the various cytochromes P450 can result in an altered response to a drug (individuals can be hypo- or hyper-responsive), a toxic response to unusual levels of a drug or metabolite, and individual sensitivity to chemical carcinogens.
Multiple Drug Resistance
Cancer cells that become resistant to one chemotherapeutic drug often become resistant to an entire group of chemotherapeutic drugs. This phenomenon is usually called multiple drug resistance (MDR).
Many patients on chemotherapy initially have striking remissions but later relapse and die from cancer that exhibits resistance to a wide array of structurally unrelated antineoplastic agents. The MDR phenomenon includes cross-resistance among the anthracyclines, the epipodophyllotoxins, the vinca alkaloids, taxol, and other compounds. A number of drugs are able to reverse MDR, including calcium channel blockers, phenothiazines, quinidine, antimalarial agents, antiestrogenic and other steroids, and cyclosporine. Liposome therapy also reverses MDR, with or without a drug on board.
In vitro studies in the past indicate that this form of resistance is associated with amplification or over-expression of the mdr-1 gene in tumors. The mdr-1 gene codes for the expression of a cell surface protein, P-glycoprotein (P-gp), a transmembrane protein which acts as an energy-dependent efflux pump that transports drugs associated with MDR out of the tumor cell before cytotoxic effects occur. ATP hydrolysis on the cytoplasmic face of P-gp is required for export of hydrophobic compounds from a tumor cell.
Normal mdr-1 expression occurs in secretory epithelial cells of the liver, pancreas, small intestine, colon, and kidney; endothelial capillary cells of the brain and testis; placental trophoblasts; and the adrenal gland. In the liver, P-gp is localized on the biliary domain of the hepatocyte membrane. In the small intestine and colon, P-gp is present on the luminal side of epithelial cells. P-gp transports dietary toxins back into the lumen and therefore helps prevent the toxins from being absorbed into the portal circulation.
Clinical studies have also previously shown that pharmaceuticals that are effective in eliminating MDR of tumor cells in vitro (apparently by inhibition of P-gp) restore chemotherapeutic cytotoxicity in vivo. Studies with small numbers of patients suggest that the addition of verapamil, diltiazem, quinine, trifluoperazine, or cyclosporine to chemotherapeutic regimens has the potential to reverse MDR.
Absorption By The Gut
Absorption across epithelia, in particular intestinal epithelia, also affects drug bioavailability. The intestine lumen presents a convoluted surface that increases the surface area of the intestine to facilitate absorption of both nutrients and drugs. The membrane of the enterocyte contains many transport proteins that actively carry nutrients from the lumen of the gut into the interior of the enterocytes. Many molecules, including many drugs, passively diffuse or are actively transported through the membrane and into the cytoplasm. Most nutrients and drugs pass through the enterocyte and eventually diffuse into the capillary net on route to the portal circulation system and the liver.
The intestine can also pump drugs out of the intestine and back into the lumen. The ability of the intestine to pump drugs out of the tissue has been thought to be important in protection against potentially damaging hydrophobic cations and toxins and for protection against small intestine cancer. No drugs or formulations have been designed to reduce pumping of drugs back into the intestine to increase drug bioavailability prior to the present invention.
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